The membranes of neuronal and retinal tissue are high in phospholipids containing one or two long chain polyunsaturated acyl chains. Docosahexaenoic acid (abbreviated DHA or 22:6n-3) is the major polyunsaturated acyl chain in these tissues. This study is directed towards determining a molecular basis for the modulation of G protein-coupled receptor signaling by polyunsaturated phospholipids, in particular those containing 22:6n-3 acyl chains. Studies are also focused on the response of these phospholipids to the acute exposure to ethanol. Fluorescence Energy Transfer (FRET) studies have demonstrated lateral domain formation in bilayers containing di16:0PC and di22:6 PC plus cholesterol, which was dependent on the presence of both cholesterol and rhodopsin. The role of cholesterol was clarified by developing a new technique to determine the partition coefficient for cholesterol in various lipid bilayers. The results demonstrate that cholesterol has the lowest partition coefficient in lipids containing DHA side chains and the highest partitioning in saturated acyl chain lipids. Therefore, lateral domain formation is likely driven by the interaction of cholesterol with the di16:0PC acyl chains and the preferential interaction of rhodopsin with the di22:6PC. An interesting result of the partitioning studies is that cholesterol partitions to a much greater degree into lipid bilayers containing trans fatty acids than cis fatty acids. Incorporation of trans fatty acids in the membrane lipids of the endothelial lining of blood vessels would result in higher cholesterol levels, which may well contribute to the formation of atherosclerotic plaques. In studies of the rod outer segment disk membrane derived from control and n-3 deficient rats, we have demonstrated differences in the phospholipid acyl chain packing properties that result from the substitution of DPA, 22:5n-6, an n-6 acyl chain for the n-3 DHA acyl chain. These two side chains differ by only a single double bond, yet the substitution was found to have marked functional consequences for the efficacy of G protein-coupled signaling in the visual transduction pathway. The studies carried out here are important in developing an understanding the role lipid composition in general and 22:6n-3 acyl chains in particular, in domain formation, the modulation of membrane protein function, the heterogeneous distribution of cholesterol in cell membranes, and the health benefits and deficits associated with polyunsaturated fatty acids and trans fatty acids, respectively.